Their primary purpose is to introduce or unmask a polar functional group—such as a hydroxyl (-OH), amino (-NH₂), sulfhydryl (-SH), or carboxyl (-COOH) group—onto the parent drug molecule.

• This modification is achieved through chemical reactions of oxidation, reduction, or hydrolysis.

• The resulting metabolite is typically only slightly more water-soluble than the parent drug but now possesses a chemically reactive "handle" that makes it a suitable substrate for the next stage of metabolism.⁸

Thus, Phase I reactions modify the drug’s chemical structure by oxidation, reduction, or hydrolysis processes.

• Phase I metabolites are not necessarily inactive – some are equally or more active than the parent compound, and some can be toxic.

  • Sometimes on inactive drug is metabolized to become of the active form and the inactive drug is typically referred to as a "prodrug."¹

    • An example of a prodrug activated by the cytochrome P450 system is cyclophosphamide, an antitumor agent, forming the "cell-killing" electrophilic form.

      • Another example is the anti-thrombotic drug clopidogrel, ultimately metabolized to an irreversible inhibitor form acting at platelet ADP P2Y₁₂ receptors.¹

• Phase I metabolism is predominantly carried out by a family of oxidative enzymes known as cytochrome P450 (CYP) monooxygenases, located mainly in the smooth endoplasmic reticulum of hepatocytes.⁵

  • The CYP450 system (also called the microsomal mixed-function oxidase system) is responsible for the oxidation of a wide variety of drugs. Key CYP isoenzymes include CYP3A4/5, CYP2D6, CYP2C9, CYP2C19, CYP1A2, among others.

    Over 75% of phase I metabolism are due to the activity of the cytochrome P450 drug metabolizing system.

    • As a group these enzymes are classified as heme-coupled monooxygenases, binding oxygen (also binding carbon monoxide which leads to the wavelength absorption at 450 nm, providing the namesake for this enzyme system).

    • CYP3A4 is the most abundant isoform in the liver and alone accounts for ~50% of drug metabolism.⁵  

    • CYP2D6, CYP2C9, and CYP2C19 are also highly clinically relevant; together these enzymes metabolize a large portion of commonly used medications.

      • Non-CYP enzymes contributing to Phase I reactions include flavin-containing monooxygenases (FMOs), alcohol/aldehyde dehydrogenases, monoamine oxidases, and various esterases and amidases (which catalyze hydrolysis of ester or amide bonds).^11,12 

• Although we will be discussing phase II reactions later, it is notable that the clinician may prescribe the same dose of an agent to two people and get different outcomes as result of differing states of drug metabolizing systems.

  • For example, underlying liver disease, age extremes (neonates and the elderly), and genetic polymorphisms in metabolizing enzymes can all impact the speed at which a drug is metabolized and cleared.^13,14 

  • Elderly patients often have reduced Phase I (CYP450-mediated) metabolism, while Phase II pathways are relatively preserved.¹⁵ 

  • Newborn infants have immature Phase II conjugation capacity – a classic illustration is “gray baby syndrome” from neonatal chloramphenicol toxicity, caused by infants’ inability to glucuronidate and clear the drug.¹⁶

Cytochrome P450 Isoenzymes (Isoforms)

• Important CYP isoenzymes include CYP3A4/5, CYP2D6, CYP2C9, CYP2C19, CYP1A2, among others.5

  • Of these, CYP3A4 is the most abundant isoform in the liver, accounting for ~50% of drug metabolism.¹⁷

• Attribution: The figure above corresponds to figure 1 of reference 17. Sychev D Ashraf G Svistunov A Maksimov M Tarasov V Chubarev V Otdelenov B Denisenko N Barreto G Alieve G The cytochrome P450 isoenzyme-some new opportunities for the prediction of negative drug interactions in vivo. Drug Des Devel Ther. 2018 May 8;12:1147-1156. https://pmc.ncbi.nlm.nih.gov/articles/PMC5951216/ (17)

Oxidation Reactions

• Oxidation reactions are the most common Phase I processes.

  • Oxidation reactions usually  involve inserting an oxygen atom or removing electrons/hydrogens from the drug.¹⁸ 

    • The CYP450 enzymes catalyze oxidation via a complex cycle requiring NADPH and O_{2 .}

    • Overall, Phase I reaction using the CYP450 system includes both an oxidative and reductive step using NADPH and not ATP.⁵

      • Drug + O₂+ NADPH → Drug*+ H₂0 + NADP+.⁵

  • Examples

    • Metoprolol¹⁹  (a β-adrenergic receptor antagonist) and omeprazole²¹   (a proton pump inhibitor) are oxidatively metabolized by CYP2D6 and CYP2C19, respectively.^19,21 

      • Enzyme polymorphism can result in markedly different drug levels in patients (poor vs. ultrarapid metabolizers).^20,21

    • Codeine is a classic example of an activation via a Phase I reaction.

      • Codeine is a prodrug that is oxidatively O-demethylated by CYP2D6 into morphine, which is the active opioid.

        • Patients who are CYP2D6 poor metabolizers derive little analgesic effect from codeine.

        • By contrast, ultrarapid metabolizers convert codeine to morphine quickly and abundantly, risking morphine toxicity (e.g. excessive CNS depression) even at standard doses.²²

          • Explanation:²³ 

            • A patient with two nonfunctional alleles of the CYP2D6 subfamily would be expected to exhibit limited drug metabolism.

            • An individual with one or two functional alleles would exhibit extensive codeine metabolism.

            • Further, someone exhibiting duplicated or amplified active CYP2D6 genes would be classified as having ultrarapid metabolism.

              •  Reduced activity of CYP3A4 (e.g. by other medications or reduced renal function) which contribute to codeine toxicity.

              • "Metabolic pathways of Codeine Biotransformation"²³

                Attribution: "The conversion of codeine into norcodeine by CYP3A4 and into codeine-6-glucuronide usually represents 80% of codeine clearance, and conversion of codeine into morphine by CYP2D6 represents only 10% of codeine clearance (Blue arrows). "Morphine is further metabolized into morphine-6-glucuronide and into morphine-3-glucuronide. Morphine and morphine-6-glucuronide have opioid activity. (Green arrows). "Glucuronides are eliminated by the kidney and are thus susceptible to accumulation cases acute renal failure. "The patient (Red arrows) had ultrarapid CYP2D6 metabolism, inhibition of CYP3A4 as result of treatment with clarithromycin and voriconazole, and glucuronide accumulation due to acute renal failure. Red arrows with dotted lines indicate low levels of drug conversion or elimination, green arrows with dotted lines indicates low levels of brain penetration, and thick arrows indicate high levels." Figure 1 from Reference 23 Gasche Y Daali Y Fathi M Chiappe A Cottini S Dayer P Desmeules J Codeine Intoxication Associated with Ultrarapid CYP2D6 Metabolism .N. Eng J Med 2004;351: 2827- 2031. https://www.nejm.org/doi/10.1056/NEJMoa041888 (Reference 23)

     

    • Another example is the antiplatelet prodrug clopidogrel, which requires CYP2C19-mediated oxidation to form its active thiol metabolite.

      • Individuals with loss-of-function CYP2C19 alleles (≈2% of Caucasians, up to 15–20% of Asians) are “poor metabolizers” who activate clopidogrel poorly.

        • These patients will have a reduced antiplatelet effect and are at higher risk of stent thrombosis; the FDA label for clopidogrel carries a warning about diminished efficacy in CYP2C19 poor metabolizers and suggests considering alternative antiplatelet agents in such cases.²⁴

    • Phase I oxidation can sometimes produce a toxic metabolite

      • The analgesic acetaminophen (paracetamol) is mainly cleared by Phase II conjugation, but about 5–10% of a therapeutic dose is oxidized by CYP2E1 and other CYPs to a reactive intermediate NAPQI (N-acetyl-p-benzoquinone imine).

        • NAPQI is normally detoxified by conjugation with glutathione.

        • In acetaminophen overdose, however, the primary sulfation and glucuronidation pathways saturate and more drug undergoes Phase I oxidation.

          • Excessive NAPQI is generated, which depletes hepatic glutathione and then covalently binds to liver proteins, causing hepatocellular necrosis.²⁵

            • Administering (N-acetylcysteine) Increases cellular glutathione levels which in turn promotes NAPQI conjugation that reduces toxicity.²⁶

             

• Attribution: The figure above corresponds to figure 1 of reference 17. Sychev D Ashraf G Svistunov A Maksimov M Tarasov V Chubarev V Otdelenov B Denisenko N Barreto G Alieve G The cytochrome P450 isoenzyme-some new opportunities for the prediction of negative drug interactions in vivo. Drug Des Devel Ther. 2018 May 8;12:1147-1156. https://pmc.ncbi.nlm.nih.gov/articles/PMC5951216/ (17)